The acute respiratory distress syndrome (ARDS) is a severe (>30% mortality) critical illness that affects over 190,000 patients in the United States each year. Despite over 40 years of study, little is known about how this disease develops, and no specific treatment exists. While sepsis is a well-known cause of ARDS, the mechanisms underlying the development of septic ARDS are uncertain. A key step in the development of ARDS is dysfunction of the pulmonary endothelial barrier, which separates blood from the lung interstitium. Studies of the systemic (non-pulmonary) vasculature have revealed that proper endothelial barrier function is dependent on an intact glycocalyx-a thin layer of proteoglycans, glycoproteins, and glycosaminoglycans lining the vascular lumen. Emerging data suggest that inflammatory mediators of sepsis degrade the systemic endothelial glycocalyx, causing barrier dysfunction. The role of the pulmonary endothelial glycocalyx in sepsis and ARDS, however, has been unexplored. We hypothesize that sepsis induces pulmonary endothelial glycocalyx degradation, leading to barrier dysfunction and ARDS. As glycocalyx structure is often aberrant in-vitro, this hypothesis will be explored using ex-vivo and in-vivo models: the isolated, perfused mouse lung and closed-chest, intravital mouse lung microscopy. The dose-response and time course of lipopolysaccharide (LPS, a model of sepsis) induced pulmonary glycocalyx degradation will be determined. The vascular segmental (arterial vs. microvascular vs. venous) pattern of glycocalyx loss will be mapped using multiphoton intravital microscopy and compared to the segmental pattern of endothelial hyperpermeability during sepsis. The role of tumor necrosis factor 1 and heparanase will be explored as mediators of LPS-induced glycocalyx loss, using pharmacologic inhibitors and transgenic mouse models. The therapeutic benefit of heparin, an inhibitor of heparanase, will be explored. In summary, this investigation promises a better understanding of a poorly-understood structure (the pulmonary endothelial glycocalyx) with relevance to a common clinical condition (sepsis-associated ARDS) via a combined physiologic (isolated, perfused mouse lung) and specialized microscopic (multiphoton intravital microscopy) approach. Furthermore, this investigation identifies heparanase as an attractive therapeutic target in a disease that yet lacks specific treatment. PUBLIC HEALTH RELEVANCE: Sepsis is a common, severe condition caused by the body's response to an overwhelming infection. Sepsis often causes blood vessels in the lungs to become leaky, flooding the lungs with blood and fluid (an often-fatal condition called the acute respiratory distress syndrome or ARDS). We are studying how to prevent ARDS, specifically by trying to protect a thin, yet very important, layer of sugars lining the blood vessels in the lung.